Inhibition of energy-producing pathways of HepG2 cells by 3-bromopyruvate.
3-BrPA (3-bromopyruvate) is an alkylating agent with anti-tumoral activity on hepatocellular carcinoma. This compound inhibits cellular ATP production owing to its action on glycolysis and oxidative phosphorylation; however, the specific metabolic steps and mechanisms of 3-BrPA action in human hepatocellular carcinomas, particularly its effects on mitochondrial energetics, are poorly understood. In the present study it was found that incubation of HepG2 cells with a low concentration of 3-BrPA for a short period (150 microM for 30 min) significantly affected both glycolysis and mitochondrial respiratory functions. The activity of mitochondrial hexokinase was not inhibited by 150 microM 3-BrPA, but this concentration caused more than 70% inhibition of GAPDH (glyceraldehyde-3-phosphate dehydrogenase) and 3-phosphoglycerate kinase activities. Additionally, 3-BrPA treatment significantly impaired lactate production by HepG2 cells, even when glucose was withdrawn from the incubation medium. Oxygen consumption of HepG2 cells supported by either pyruvate/malate or succinate was inhibited when cells were pre-incubated with 3-BrPA in glucose-free medium. On the other hand, when cells were pre-incubated in glucose-supplemented medium, oxygen consumption was affected only when succinate was used as the oxidizable substrate. An increase in oligomycin-independent respiration was observed in HepG2 cells treated with 3-BrPA only when incubated in glucose-supplemented medium, indicating that 3-BrPA induces mitochondrial proton leakage as well as blocking the electron transport system. The activity of succinate dehydrogenase was inhibited by 70% by 3-BrPA treatment. These results suggest that the combined action of 3-BrPA on succinate dehydrogenase and on glycolysis, inhibiting steps downstream of the phosphorylation of glucose, play an important role in HepG2 cell death.
Pereira da Silva
Ana Paula
AP
Laboratório de Bioenergética e Fisiologia Mitocondrial, Programa de Bioquímica e Biofísica Celular, Instituto de Bioquímica Médica, Universidade Federal do Rio de Janeiro, Av. Carlos Chagas Filho 373-CCS, Bl. D, ss13, 21941-902, RJ, Brazil.
El-Bacha
Tatiana
T
Kyaw
Nattascha
N
dos Santos
Reinaldo Sousa
RS
da-Silva
Wagner Seixas
WS
Almeida
Fabio C L
FC
Da Poian
Andrea T
AT
Galina
Antonio
A
D000255:Adenosine Triphosphate / Q000037:antagonists & inhibitors* / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000378:metabolism*; D004734:Energy Metabolism / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000037:antagonists & inhibitors / Q000378:metabolism; D006019:Glycolysis / Q000187:drug effects; D006801:Humans; D019344:Lactic Acid / Q000378:metabolism; D008113:Liver Neoplasms / Q000378:metabolism*; D008928:Mitochondria / Q000187:drug effects*; D010085:Oxidative Phosphorylation / Q000187:drug effects; D010101:Oxygen Consumption / Q000187:drug effects; D011773:Pyruvates / Q000494:pharmacology*; D013385:Succinate Dehydrogenase / Q000037:antagonists & inhibitors / Q000378:metabolism
D004791:Enzyme Inhibitors; D011773:Pyruvates; D019344:Lactic Acid; C017092:bromopyruvate; D000255:Adenosine Triphosphate; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D013385:Succinate Dehydrogenase